Cell sorting system using microfabricated components

ABSTRACT

A MEMS-based cell sorter is disclosed, which uses a novel combination of features to accomplish the cell sorting using a microfabricated cell sorting valve housed in a disposable cartridge. The features include an interposer that provides fluid communication between the microfluidic passages in the silicon substrate and a plurality of fluid reservoirs in the cartridge, including a sample, sort and waste reservoir The disposable cartridge may include other features that assist in the handling of small volumes of fluids, such as a siphon region in the sort reservoir and funnel-shaped regions in the sample and waste reservoirs. A mixing mechanism may be provided for stirring the contents of the sample reservoir.

CROSS REFERENCE TO RELATED APPLICATIONS

This U.S. Patent Application is a continuation-in-part of U.S. patentapplication Ser. No. 13/998,095, filed Oct. 1, 2013, which isincorporated by reference herein in its entirety.

STATEMENT REGARDING FEDERALLY SPONSORED RESEARCH

Not applicable.

STATEMENT REGARDING MICROFICHE APPENDIX

Not applicable.

BACKGROUND

This invention relates to a cell sorting system using a microfabricated,movable cell sorting mechanism.

Microelectromechanical systems (MEMS) are very small, often moveablestructures made on a substrate using surface or bulk lithographicprocessing techniques, such as those used to manufacture semiconductordevices. MEMS devices may be moveable actuators, sensors, valves,pistons, or switches, for example, with characteristic dimensions of afew microns to hundreds of microns. A moveable MEMS switch, for example,may be used to connect one or more input terminals to one or more outputterminals, all microfabricated on a substrate. The actuation means forthe moveable switch may be thermal, piezoelectric, electrostatic, ormagnetic, for example. MEMS devices can also be made which manipulateparticles passing by the MEMS device in a fluid stream.

For example, a MEMS device may be a movable valve, used as a sortingmechanism for sorting various particles from a fluid stream, such ascells from blood. The particles may be transported to the sorting devicewithin the fluid stream enclosed in a microchannel, which flows underpressure. Upon reaching the MEMS sorting device, the sorting devicedirects the particles of interest such as a blood stem cell, to aseparate receptacle, and directs the remainder of the fluid stream to awaste receptacle.

Previously, particle sorters existed using fluorescence-activated cellsorting (FACS) and are known as flow cytometers. Flow cytometers aregenerally large and expensive systems which sort cells based on afluorescence signal from a tag affixed to the cell of interest. Thecells are diluted and suspended in a sheath fluid, and then separatedinto individual droplets via rapid decompression through a nozzle. Afterejection from a nozzle, the droplets are separated into different binselectrostatically, based on the fluorescence signal from the tag. Amongthe issues with these systems are cell damage or loss of functionalitydue to the decompression, difficult and costly sterilization proceduresbetween samples, inability to re-sort sub-populations along differentparameters, and substantial training necessary to own, operate andmaintain these large, expensive pieces of equipment. For at least thesereasons, use of flow cytometers has been restricted to large hospitalsand laboratories and the technology has not been accessible to smallerentities.

MEMS-based cell sorting systems may have substantial advantages overflow cytometers in terms of cost, speed and size. A number of patentshave been granted which are directed to such MEMS-based particle sortingdevices. For example, U.S. Pat. No. 6,838,056 (the '056 patent) isdirected to a MEMS-based cell sorting device, U.S. Pat. No. 7,264,972 b2(the '972 patent) is directed to a micromechanical actuator for aMEMS-based cell sorting device. U.S. Pat. No. 7,220,594 (the '594patent) is directed to optical structures fabricated with a MEMS cellsorting apparatus, and U.S. Pat. No. 7,229,838 (the '838 patent) isdirected to an actuation mechanism for operating a MEMS-based particlesorting system. Additionally, U.S. patent application Ser. No.13/374,899 (the '899 application) and Ser. No. 13/374,898 (the '898application) provide further details of other MEMS designs. Each ofthese patents ('056, '972, '594 and '838) and patent applications ('898and '899) is hereby incorporated by reference.

Among the problems encountered with using microfluidic devices in thecell sorting systems as mentioned above, is the clogging of the narrowpassageways, and the interface of these narrow passageways with themacroscopic world, and control of the movement of these very small,movable devices.

SUMMARY

A cell sorting system is described which makes use of a microfabricatedcell sorting MEMS chip. The passageways in the MEMS chip are formedlithographically, and are thus very small. Clogging of these narrowpassageways presents a significant challenge to reliable, long termoperation. Additionally, these narrow passageways must be mated to muchlarger, macroscopic features, and handle small volumes of fluids,particularly when sorting rare cells.

In the system described here, various novel design elements are broughtto bear to enable such a MEMS cell sorting system. The sorting mechanismmay be a MEMS fluid valve formed on a silicon substrate, which may beadhered to an interposer and installed in a disposable cartridge. Thecartridge may provide all of the fluidic passageways for the handling ofthe sample fluid, and may include larger reservoirs (e.g. sort, sampleand waste reservoirs) for the storage of volumes of fluids. A plasticinterposer may be used to provide the interconnections between themicroscopic passages of the MEMS fluid valve and the macroscopicfeatures of the reservoirs. The MEMS fluid valve, interposer andreservoirs may all be contained in a disposable cartridge, such thatsterilization of the cell sorting system is straightforward, thecartridge is simply disposed of.

A specially designed electromagnet may provide the precisely locatedelectromagnetic fields which cause the very small MEMS chip to movewithin the much larger system. This electromagnet minimizes heatproduced, and thus improves efficiency. Finally, a special formulationof fluid materials is used to reduce or eliminate clogging.

The disposable cartridge and interposer may include a number of novelfeatures, such a s a mixer, and funnel-shaped regions that may assist inthe handling of small volumes of fluids. The mixer may be submerged inthe sample reservoir, thereby allowing mixing of the contents.Funnel-shaped regions may be provided in the sort reservoir, the samplereservoir, and the waste reservoir, for the collection of small volumesof fluids.

Accordingly, a cell sorting system is described, which may include acell sorting valve microfabricated on a silicon substrate withmicrofabricated channels leading from the cell sorting valve, adisposable cartridge containing a sample reservoir, a sort reservoir anda waste reservoir and an interposer that that provides fluidcommunication between the microfabricated channels in the siliconsubstrate and the reservoirs in the disposable cartridge. The cellsorting system may further include an electromagnet with a tapered tip,coils and magnetic core, wherein the tapered shape serves to concentratethe lines of flux produced by the coils and core, and exit from theelectromagnet in the vicinity of the tip. Finally, a cation-independentDNAse may be used as a buffer in which to suspend the target particles,reducing or eliminating clogging of the small channels in the device.

BRIEF DESCRIPTION OF THE DRAWINGS

Various exemplary details are described with reference to the followingfigures, wherein:

FIG. 1 is a schematic illustration of a MEMS chip sorter in a firstposition;

FIG. 2 is a schematic illustration of a MEMS chip sorter in a secondposition;

FIG. 3 is a schematic illustration of an exemplary MEMS cell sortingsystem which may make use of the MEMS sorter of FIGS. 1 and 2;

FIG. 4 is an exploded view of an exemplary disposable cartridge whichmay be used in the MEMS cell sorting system of FIG. 3, which includes aMEMS chip sorter and an interposer;

FIG. 5a is a side view of the exemplary disposable cartridge which maybe used in the MEMS cell sorting system of FIG. 3, which includes a MEMSchip sorter and an interposer; FIG. 5b is an end view of the exemplarydisposable cartridge;

FIG. 6 is another side view of the exemplary disposable cartridge whichmay be used in the MEMS cell sorting system of FIG. 3, which includes aMEMS chip sorter and an interposer

FIG. 7 is a plan view of an exemplary interposer which may be used withthe disposable cartridge of FIG. 4;

FIG. 8a is a perspective view of the exemplary interposer which may beused with the disposable cartridge of FIG. 4, showing thecartridge-facing side; FIG. 8b is a cross sectional view of a channel;FIG. 8c is a cross sectional illustration of another embodiment of achannel;

FIG. 9 is a perspective view of the obverse side of the exemplaryinterposer;

FIG. 10 is a perspective view of the obverse side of the exemplaryinterposer showing calibration areas;

FIG. 11 is a cross sectional view of the interposer with calibrationareas; and

FIG. 12a is a plan view of the targeted electromagnet that may generatethe magnetic field which may actuate the MEMS chip sorter from the firstposition (FIG. 1) to the second position (FIG. 2); FIG. 12b is a closeupview of the magnet tip; FIG. 12c is a perspective view of the targetedelectromagnet;

It should be understood that the drawings are not necessarily to scale,and that like numbers may refer to like features.

DETAILED DESCRIPTION

Systems and methods are described for sorting target particles fromnon-target materials in a fluid stream. The systems and methods make useof a microfabricated (MEMS) movable valve or sorting mechanism, whichdirects the target particle from a sample input passageway into a sortpassageway, while allowing non-target material to flow into a wastepassageway. Both the sort and waste passageways lead to a separate,respective reservoir, the sort and the waste reservoir, and are storedthere until removal. The sort, sample and waste reservoirs, along withthe MEMS chip sorter, may be contained in a plastic disposablecartridge. This cartridge may then be discarded after the fluids arecollected from the reservoirs. This allows greatly reduced burden forsterilizing the system between samples. The systems and methods may alsohave significant advantages in terms of cost, performance, speed andcomplexity. The system may also be substantially gentler in its handlingof cells, such that viability of cells in the effluent is greatlyimproved compared to droplet-based flow cytometers.

Because of the microfluidic nature of this cell sorting system, measuresare taken to reduce or eliminate clogging, and to handle the smallvolumes of fluids, and to control the very small movable valve. Aninterposer may be used to provide the interconnections between themicroscopic passages and the macroscopic features. Finally, speciallydesigned electromagnet provides the precisely located electromagneticfields which cause the very small MEMS chip to move within the muchlarger system. This electromagnet minimizes heat produced, and thusimproves efficiency. Each of these features is described further below.

FIG. 1 is a schematic diagram of a microfabricated cell sortingmechanism, MEMS chip sorter 10, which may be used in the particlesorting system described here. Details of cell sorting mechanism may befound in co-pending U.S. patent application Ser. No. 13/998,095,(hereinafter the '095 patent application) incorporated by referenceherein. Among the unique features of microfabricated cell sortingmechanism 10 is that the motion of the cell sorting valve 10 is parallelto the fabrication plane of the valve. In addition, the waste channel140 is substantially orthogonal to the sample inlet channel 120 and thesort output channel 122. These features enable distinct advantages interms of speed and precision, valve throughput and ease of themicrofluidic sorting.

It should be understood that the term “chip sorter” 10 is an abbreviatedterm for a microfabricated cell sorting valve 10, as a “chip” is adevice microfabrucated on a substrate. Either term is intended todesignate a microfabricated movable valve formed on a surface of asubstrate, using MEMS fabrication techniques, which, by its movement, iscapable of separating target particles from non-target material.

In the plan view illustration of FIG. 1, the novel MEMS chip sorter orcell sorting valve 10 is in the quiescent (un-actuated) position. Thechip sorter 10 may include a microfabricated fluidic valve or movablemember 110 (hatched area) and a number of microfabricated fluidicchannels 120, 122 and 140. Microfabricated fluidic channel 140 (shown asdashed area 140 in FIG. 1 and FIG. 2) serves as output channel and ismay be located directly below at least a portion of the microfabricatedmember 110 and is not parallel to the plane of the microfabricatedfluidic channels 120, 122 or the microfabricated member 110.Microfabricated member 110 is fabricated and moves in a path parallel orwithin this plane. Preferably, the microfabricated fluidic channel 140is orthogonal to the plane of the microfabricated fluidic channels 120,122 and the path of motion of microfabricated member 110. The apertureof microfabricated fluidic channel 140 may cover preferably overlap atleast a portion of the path of motion of microfabricated member 110,i.e. the dashed, area overlaps the microfabricated member 110 over atleast a portion of its motion, as shown in FIG. 1 and FIG. 2. Thisoverlap may allow a fluid path to exist between the input channel 120and the output channel 140 when the microfabricated member is in the“waste” or unactuacted position (FIG. 1), and this path is closed offand the particles redirected in the “sort” or actuated position (FIG.2). As described previously, this architecture may reduce the fluidresistance, thereby increasing the speed of microfabricated member 110.

The movable member 110 and microfabricated fluidic channels 120, 122 and140 may be formed on the surface of a suitable substrate, such as asilicon substrate, using MEMS lithographic fabrication techniques asdescribed in greater detail in the '095 application. The fabricationsubstrate may have a fabrication plane in which the device is formed andin which the movable member 110 moves.

A sample stream may be introduced to the microfabricated movable member110 by a sample inlet channel 120. The sample fluid may be stored in asample reservoir 20 prior to sorting by movable member 110. The samplestream may contain a mixture of particles, including at least onedesired, target particle and a number of other undesired, non-targetparticles. The particles may be suspended in a fluid. For example, thetarget particle may be a biological material such as a stem cell, acancer cell, a zygote, a protein, a T-cell, a bacteria, a component ofblood, a DNA fragment, for example, suspended in a buffer fluid such assaline, or the novel chemistry described below. The inlet channel 120may be formed in the same fabrication plane as the movable member 110,such that the flow of the fluid is substantially in that plane. Themotion of the cell sorting valve 10 is also within this fabricationplane. The decision to sort/save or dispose/waste a given particle maybe based on any number of distinguishing signals. In one exemplaryembodiment, the decision is based on a fluorescence signal emitted bythe particle, based on a fluorescent tag affixed to the particle andexcited by an illuminating laser. Laser interrogation region 200 is theportion of the microfluidic passageway in which an illuminating orinterrogating laser is directed on the target particle, in order todistinguish it from the other constituents of the fluid sample. Detailsas to this detection mechanism are well known in the literature, andfurther discussed below with respect to FIG. 3. However, other sorts ofdistinguishing signals may be anticipated, including scattered light orside scattered light which may be based on the morphology of a particle,or any number of mechanical, chemical, electric or magnetic effects thatcan identify a particle as being either a target particle, and thussorted or saved, or an non-target particle and thus rejected orotherwise disposed of.

With the movable member 110 in the position shown, the input streampasses unimpeded to a waste outputchannel 140 which is out of the planeof the inlet channel 120, and thus out of the fabrication plane of theMEMS chip sorter 10. That is, the flow is from the inlet channel 120 tothe output orifice 140, from which it flows substantially vertically,and thus orthogonally with respect to the inlet channel 120. This outputorifice 140 leads to an out-of-plane channel that may be perpendicularto the plane of the paper showing FIG. 1. More generally, the wasteoutput channel 140 is not parallel to at least one of the plane of theinlet channel 120 or sort channel 122, or the fabrication plane of themovable member 110. In one embodiment, the sort and sample channels maybe antiparallel, that is, flow in the sort channel is in an oppositedirection to flow in the incoming sample channel.

Accordingly, the cell sorting system may include a cell sorting valve10, which directs the target particles from a sample channel 120 into asort channel 122 formed in the silicon substrate and the non-targetmaterial from the sample channel 120 to a waste output channel 140 alsoformed in the silicon substrate. The cell sorting valve 10 may also movein a plane parallel to the surface, and direct the target particles fromthe sample channel 120 into the waste channel 140 when themicrofabricated cell sorting valve 10 is in a first position, and whichdirects the other particles into the sort channel 122 when in a secondposition, wherein the sort channel 122 and the waste channel 140 aresubstantially antiparallel, and the sample channel 120 and waste channel140 are substantially orthogonal.

The waste output channel 140 may have an orifice, which may be a holeformed in the fabrication substrate, or in a covering substrate that isbonded to the fabrication substrate. Further, the movable member 110 mayhave a curved diverting surface 112 which can redirect the flow of theinput stream into a sort output stream. The contour of the surface 112may be such that redirects the sample stream from the inlet channel 120into the sort channel 122 in one position, while allowing it to flow tothe waste output channel 140 in another position. Accordingly, by havingthe surface 112 overlap the inlet channel 120, a route exists for theinput stream to flow directly into the waste output channel 140 when themovable member 110 is in the un-actuated waste position, as is shown inFIG. 1. The waste output channel 140 may lead to a waste reservoir 40,which may collect the non-target material. The inlaid magneticallypermeable material 116 on the movable member 110 may cause its movement,and will be described below in the description of FIG. 2.

FIG. 2 is a plan view of the MEMS chip sorter 10 in the actuatedposition. In this position, the movable member 110 or valve 10 isdeflected upward into the position shown in FIG. 2. The curved divertingsurface 112 is a sorting contour which redirects the flow of the inletchannel 120 into the sort output channel 122. The output channel 122 maylie in substantially the same plane as the inlet channel 120, such thatthe flow within the sort channel 122 is also in substantially the sameplane as the flow within the inlet channel 120. There may be an angle □between the inlet channel 120 and the sort channel 122. This angle maybe any value up to about 90 degrees. Actuation of movable member 110 mayarise from a force from force-generating apparatus 400, showngenerically in FIG. 2. In some embodiments, force-generating apparatus400 may be an electromagnet, as described above. However, it should beunderstood that force-generating apparatus may also be electrostatic,piezoelectric, or some other means to exert a force on movable member110, causing it to move from a first position (FIG. 1) to a secondposition (FIG. 2). The sort channel 122 may lead to a sort reservoir 22which collects the sorted, target particles as effluent from the movablevalve in the position shown in FIG. 2. The inlet channel 120 may conductthe sample fluid from an sample reservoir 20 to the waste channel 140and waste reservoir 40 as was shown in FIG. 1, or to the sort channel122 and sort reservoir 22, as shown in FIG. 2.

In some embodiments, the force generating apparatus 400 may includecoils which generate a magnetic field, which then interacts with themovable member 110. In order to make the movable member 110 responsiveto such an electromagnetic force, it may have a magnetically permeablematerial inlaid into movable valve 110. The extent of this inlaidmagnetic material 116 may be just inside the edge of the outline of themovable member 110 as shown by the dashed lines in FIGS. 1 and 2.

A magnetically permeable material should be understood to mean anymaterial which is capable of supporting the formation of a magneticfield within itself. In other words, the permeability of a material isthe degree of magnetization that the material obtains in response to anapplied magnetic field.

The terms “permeable material” or “material with high magneticpermeability” as used herein should be understood to be a material witha permeability which is large compared to the permeability of air orvacuum. That is, a permeable material or material with high magneticpermeability is a material with a relative permeability (compared to airor vacuum) of at least about 100, that is, 100 times the permeability ofair or vacuum which is about 1.26×10⁻⁶ H·m⁻¹. There are many examples ofpermeable materials, including chromium (Cr), cobalt (Co), nickel (Ni)and iron (Fe) alloys. One popular permeable material is known asPermalloy, which has a composition of between about 60% and about 90% Niand 40% and 10% iron. The most common composition is 80% Ni and 20% Fe,which has a relative permeability of about 8,000. Accordingly, movablemember 110 may have permalloy material inlaid 116 into the movablemember 110 and subsequently planarized so that the profile of themovable valve remains flat. Additional details as to the fabrication ofsuch permeable features may be found in the incorporated '095 patentapplication.

It is well known from magnetostatics that permeable materials are drawninto areas wherein the lines of magnetic flux are concentrated, in orderto lower the reluctance of the path provided by the permeable materialto the flux. Accordingly, a gradient in the magnetic field urges themotion of the movable member 110 because of the presence of inlaidpermeable material 116, towards areas having a high concentration ofmagnetic flux. That is, the movable member 110 with inlaid permeablematerial 116 will be drawn in the direction of positive gradient inmagnetic flux. A novel core design is described below with respect toFIG. 10a-10c , which concentrates the lines of flux in a very specificarea, to optimize the control over the movable member 110.

It should be understood that the magnetostatic embodiment describedabove is but one of a number of actuation mechanisms that can be used tomove the cell sorting valve or chip sorter 10. More generally, the cellsorting system may be constructed with a cell sorting valve 10, whereinwhen the cell sorting valve 10 is in a first position, a passage betweenthe sample channel 120 and the waste channel 140 is formed. When thecell sorting valve 10 is in the second position, a passage between thesample channel 120 and the sort channel 122 is formed. The cell sortingvalve 10 may move from the first position to the second position inresponse to the application of a force, and that force may be at leastone of mechanical, electrostatic, magnetostatic, piezoelectric andelectromagnetic. In the electrostatic embodiment, a permeable magneticmaterial is inlaid in the movable member of the microfabricated cellsorting valve 10, and a source of magnetic flux 400 is provided. Themagnetic flux interacts with the inlaid permeable magnetic material 116to move the microfabricated cell sorting valve 10, whereby themicrofabricated cell sorting valve moves from the first position to thesecond position when the source of magnetic flux 400 is activated.

FIG. 3 is a schematic illustration of the cell sorting system 1 whichmay use microfluidic passageways, a MEMS chip sorter 10 housed in adisposable cartridge 1000, and a flux-generating apparatus 400. Whatfollows is a description of some other components of the system and howthey interact with the MEMS chip sorter 10. In particular, FIG. 3 laysout the optical path of the interrogating laser for interrogation region200, and the control of fluid flow in channels 120-140 and control ofMEMS chip sorter 10. After the system level description, the discussionwill turn to the unique features of system 1 that allow the microfluidicsystem 1 to work in a precise, reliable and predictable way.

As shown in FIG. 3, the microfabricated MEMS chip sorter 10 may behoused in a disposable cartridge 1000, which may be loaded onto amovable stage and oriented with respect to detection optics 2100 andinterrogating lasers 2400 in the cell sorting system 1. Fluid then flowsthrough the MEMS chip sorter 10 from fluid reservoirs also housed indisposable cartridge 1000 through a series of passageways as will bedescribed below with respect to FIGS. 4-9.

In the normal operation of system 1, the target particle may be aparticular cell, such as a stem cell, or a cancer cell, which has beentagged with a fluorescent marker. This marker emits photons having aparticular energy when irradiated with a laser 2400 operating at apredefined wavelength. Accordingly, in this cell sorting system, a lasersource 2400 may be directed by a turning mirror 2250 through thedetection/collection optics 2100 to the laser interrogation region 200that was shown in FIGS. 1 and 2. The optical axis of thedetection/collection optics 2100 and the laser source 2400 may becollinear, at least over a portion of the optical path. Thus, theorientation of the laser application and optical detection along thisoptical axis may be perpendicular or orthogonal to the substratefabrication plane, orthogonal to the plane of motion of the movablevalve 110 and orthogonal to the flow of the sample fluid through thedetection region.

The fluorescence emitted from the irradiated particles may be shaped bydetection/collection optics 2100 and separated by dichroic mirrors 2200and directed into a bank of photodetectors 2300. A plurality ofphotodetectors may accommodate multiple wavelengths of emitted light,for multiparametric detection. The signal output by the photodetectors2300 indicates the presence or absence of the target particle in thelaser interrogation region 200. The signal may be delivered to acontroller 2900, which manages the relative timing of the components inthe particle sorting system 1, and collects the data. The controller2900 may be a general purpose computer or a specialized circuit or ASIC.Upon detection of the target particle, a signal is generated by thecontroller 2900 which energizes the force-generating or flux-generatingapparatus 400. The controller 2900 may also provide the fluidic controlto the MEMS chip sorter 10, via one or more pneumatic, hydraulic,piston-based or mechanical force-based mechanisms which are illustratedgenerically by fluid control means 2500. The rate at which particles aredetected may be monitored by the controller 2900, which may maintain thefluid control means 2500.

The force generating apparatus 400 is a device which causes a force toarise in the movable member 110 itself, causing the motion of themovable member. This force-generating apparatus 400 may not be directlymechanically coupled to the MEMS particle manipulation device 10, asindicated by the dashed line in FIG. 3. For example, theforce-generating apparatus 400 may be a source of magnetic flux whichcauses a magnetostatic force to arise in an inlaid permeable material116 in the MEMS movable member 110 of the cell sorting valve 10 asdescribed previously. Accordingly, flux generating apparatus 400 may bean electromagnet with a magnetic core and windings. This force may pullthe movable member 110 toward the force-generating apparatus 400,opening the sort channel 122 and closing the waste channel 140, as wasshown in FIGS. 1 and 2. Importantly, the force-generating apparatus 400may reside in the particle sorting system 1, rather than in the MEMSchip sorter 10. As mentioned previously, this may reduce the cost andcomplexity of the MEMS chip sorter 10, which may be housed in thedisposable portion 1000 of the system 1. In the compact system shown inFIG. 3, it is important that excessive heat not be generated byforce-generating apparatus 400. As mentioned previously, because of thevery small size of MEMS chip sorter 10, force-generating apparatus 400may also need to generate lines of flux which are concentrated in asmall area. Details as to the design of a novel flux-generatingapparatus 400 which may be suitable in this application are discussedbelow with respect to FIGS. 10a -10 c.

Another optional laser 2410 may also be included to provide a secondoptical channel in cell sorting system 1.

As mentioned, laser interrogation region 200 is the portion of themicrofluidic passageway in which the laser 2400 is directed on thetarget particle, in order to distinguish it from the other constituentsof the fluid sample.

Upon passing through the detection region 200, a signal is generated bythe detector 2300 indicating that a target particle is present in theinterrogation region 200. After a known delay, a signal is generated bythe controller 2900 which indicates that the sorting gate, i.e. themovable member 110 of the cell sorting valve 10 is to be opened, inorder to separate the target particle which was detected, from the othercomponents in the fluid stream. The movable member 110, of the MEMSvalve 10 may comprise permeable magnetic materials 116 as mentionedpreviously, so that the magnetic force may arise in it in the presenceof a magnetic field. When the signal is generated by the controller2900, a force arises in the embedded magnetically permeable material 116which draws the movable valve 110 toward the force generating apparatus400. This motion may close off waste channel 140 and redirect the targetparticle into a sort channel 122. The sorted sample is subsequentlycollected from a sort reservoir at the end of the sort channel 122,which holds the sorted sample. As mentioned previously, the controller2900 may also control flow rates based on the rate at which sortingevents are recorded.

A fluid control means 2500 may control the direction and velocity offluid flowing through the channels of the MEMS chip cell sorting valve10. The fluid control means 2500 may be controlled based on a number ofcriteria as described below. The fluid control means 2500 may includepneumatic, hydraulic, and/or one way valves, and/or may include a pistonor a pump and associated fluidic passages. During normal operation, theflow may be controlled by the fluid control means 2500 in a feedbackloop with controller 2900 to keep cell velocity, fluid pressure, orevent rate constant, for example.

In a further embodiment, the cell sorting system 1 may comprise afeedback loop to prevent clogging of the channels by cells or othersolid material suspended in the fluid. Biological cells especially tendto adhere at the channel surfaces, edges or offsets, thereby reducingthe flow of liquid through the system and/or overall cell sortingperformance. The feedback loop may consist of at least the fluid controlmeans 2500 such as a pump and the controller 2900.

The controller 2900 may detect impending clogging by monitoring thefluid pressure and/or the cell velocity within the system. If the fluidpressure and/or the cell velocity fall below a predefined threshold, itmay be indicative of impending clogging. The controller 2900 mayincrease the pump rate until the fluid pressure and/or the cell velocityreaches the threshold again. The fluid pressure can be monitored by anappropriate detector, and cell velocity can be deduced by monitoring theevent rate in the optical channel. Preferably, the cell speed may bebetween 0.2 and 10 m/s, and may be constant within +/−0.2 m/s.Accordingly the threshold activating the feedback loop may be areduction of cell speed by around 0.2 m/s or the equivalent in loss ofpressure. It should be understood that the details given here areexemplary only, and that the selection of such operating parameters willdepend on the specifics of the application.

At the end of a sorting operation when the volume of sample to be sortedin nearly exhausted, the controller 2900 in concert with the fluidcontrol means 2500 may reverse the flow of fluid in the microchannels,thus keeping the passages wet, as described in U.S. patent applicationSer. No. 14/167,566, filed Jan. 29, 2014 and incorporated by referencein its entirety. The system 1 may also have the means to evaluate theeffectiveness of the sorting process by reversing the flow through thelaser interrogation region 200, as described in detail in U.S. patentapplication Ser. No. 13/104,084, filed Dec. 12, 2013 and incorporated byreference in its entirety.

Accordingly, the cell sorting system 1 described here may include aninterrogation means 200 comprising a laser in a laser-based, inducedfluorescence system, wherein a fluorescent tag is affixed to a targetparticle, and emits a fluorescent signal when irradiated by the laser.The system may include a disposable cartridge 1000 which is configuredto be accepted into the cell sorting system 1 on a positionable stage,on which it can be positioned with respect to at least one laser source2400, and at least one optical detector 2300. The cell sorting system 1may further include a computer 2900 which is in communication with theat least one laser source 2400, at least one optical source 2100 and thecell sorting valve 10, to separate the target particles from thenon-target material.

What follows is a description of the enabling aspects of MEMS cellsorting system 1, in particular, what aspects allow the fluid to flow toand from MEMS chip sorter 10 in a repeatable and reliable way, frommacroscopic reservoirs to the MEMS chip sorter 10, and to control thevery small MEMS chip sorter 10.

FIG. 4 is an exploded perspective view of an exemplary disposablecartridge 1000 which may be used in the particle sorting system shown inFIG. 3. Disposable cartridge 1000 may include several assemblablepieces, such as top 1135 and base 1130.

Disposable cartridge 1000 may house MEMS chip sorter 10 and providestorage in fluid reservoirs. Accordingly, the base 1130 of disposablecartridge 1000 may have a plurality of voids or compartments formedtherein, including sample reservoir 20, sort reservoir 22 and wastereservoir 40. As described further below, the sample to be sorted may bestored in sample reservoir 20, the sort effluent in sort reservoir 22and waste effluent in waste reservoir 40. The fluidic passagewaysbetween these voids may all be disposed in the interposer 1400 and/or inthe MEMS chip sorter 10. Accordingly, the interposer 1400 may provide asort fluid path between a sort reservoir 22 in the disposable cartridgeand the sort channel 122 in the silicon substrate, a waste fluid pathbetween a waste reservoir 40 in the disposable cartridge and the wastechannel 140, and a sample fluid path between the sample channel 120 anda sample reservoir 20.

It should be understood that the term “sort” fluid, “sort” sample or“sort” reservoir may refer to a collection of target particles. The“waste” fluid, “waste” sample or “waste” reservoir may refer to acollection of non-target materials in the fluid stream. Other equivalentlanguage is “positive fraction” to refer to the sort sample, and“negative fraction” to refer to non-target material. Accordingly, in thetext below, “sort” portion may be equivalent to the “positive fraction”and refer to a collection of target particles, and “waste” may refer tothe negative fraction and to a collection of non-target materials.

Between the top 1135 and the base 1130 may be disposed a number offilters 1180 to protect the sample from contamination or debris. Thesefilters 1180 may be 0.20 micron Sterifilters, for example. The filters1180 may be located directly above the various fluid reservoirs 20, 22and 40. There may also be in-line filters within the fluid channels,which are for catching debris in the fluid and may be about 20 micronsin pore size.

The sample reservoir 20, sort reservoir 22 and the waste reservoir 40may also include funnel-shaped features that allow the handling of smallvolumes of fluids. The sort reservoir 22 may contain a siphon-likestructure that is described below with respect to FIG. 7, where it isillustrated in greater detail. However, the sample reservoir 20 andwaste reservoir 40 may also contain features which assist small volumehandling. This feature may be a contoured surface. A funnel-shapedfeature should be understood to mean a generally conical structure thatis shaped to collect small volumes of fluid running down the wall of thereservoir, Both the sample reservoir 20 and the waste reservoir 40 mayinclude such funnel-shaped features, 21 and 41 respectively. Thus, thecell sorting system 1 described here may include a sample reservoir 20and waste reservoir 40, wherein the sample reservoir 20 furthercomprises a funnel-shaped feature 21 formed in the wall of the samplereservoir 20, which collects smaller volumes of sample fluid, whereinthe smaller volume of sample fluid is less that about 10% of a totalfluid volume of the sample reservoir 20. Similarly, the waste reservoir40 may further comprise a funnel-shaped feature 41 formed in the wall ofthe waste reservoir 40, which collects smaller volumes of waste fluid,wherein the smaller volume of waste fluid is less that about 10% of atotal fluid volume of the waste reservoir 40.

Within the sample reservoir 20 and enclosed between the top 1135 and thebase 1130 may be a magnetized propeller 1150, and a needle 1160 whichmay act as a shaft for magnetized propeller 1150. Upon exposure to acirculating magnetic field, magnetized propeller 1150 may rotate onshaft 1160, causing the contents of the sample reservoir 20 to be mixedor homogenized. Finally, a 0.20 micron filter 1170 may be placed overthe sort reservoir 22, to protect the sorted contents from contaminationfrom the ambient environment. Alternatively, the propeller 1150 may bedriven directly by a mechanical coupling to a small motor, which maycause the rotation of the propeller 1150 and thus the mixing of thecontents of the sample reservoir 20. Details of the construction of themixing elements may be shown in more detail in FIG. 6, and discussedbelow with respect to that figure.

Sample fluid may be introduced to the sample reservoir 20 with apipette, or with a syringe and plunger (not shown) through the accessports 1111 shown, whereupon the cartridge 1000 may be sealed with maleleur lock sealing elements 1110. Alternative sealing techniques may alsobe used, such as thumbscrews. Alternatively, the cartridge 1000 may bedelivered with the sample fluid already loaded therein.

FIG. 5 is a side view of the assembled disposable cartridge 1000,showing the sample reservoir 20, sort reservoir 22 and waste reservoir40. Shown in the assembled view are the relative locations of the MEMSchip sorter 10 and interposer 1400 with respect to the cartridge base1130. It should be noted that FIG. 5 is inverted compared to FIG. 4,such that the sample reservoir 20, shown on the left hand side of thecartridge in FIG. 4, is now located on the right hand side in FIG. 5, asare the associated channels, stirrer, etc.

To provide a transition region between the very fine, microfabricatedfeatures of the MEMS chip sorter 10 and the much larger fluid volumes ofreservoirs 20, 22 and 40, an interposer 1400 may be provided. Theinterposer 1400 may be formed from plastic by, for example, injectionmolding and may have intermediate tolerances on the order of +/−10 □m.The purpose of the interposer 1400 is to provide a transition betweenthe very small structures of the MEMS device 10 and the gross,macroscopic structures of the cartridge 1000 and reservoirs 20, 22 and40. Accordingly, the cell sorting system 1 described herein may includea cell sorting valve 10 microfabricated on a surface of a siliconsubstrate, with microfabricated channels leading from the cell sortingvalve 10, wherein the cell sorting valve 10 separates the targetparticles from non-target material, a disposable cartridge 1000containing a sample reservoir 20, a sort reservoir 22 and a wastereservoir 40; and an interposer 1400 that provides fluid communicationbetween the microfabricated channels in silicon substrate and thereservoirs in the disposable cartridge.

Because the interposer 1400 can be made with reasonably fine tolerances(+/−10 □m), it is possible to align the passages in the interposer 1400with passages in the MEMS chip when the apertures to the channels are onthe order of about 300 microns. While the widths of the channels leadingto and from the movable valve 110 may be substantially smaller on theorder of 150 microns, the apertures which introduce the fluid to thechannels may be made near this scale. The apertures are shown in FIG. 6.

As shown on the insert of FIG. 6, the through holes such as 1420 ininterposer 1400 may have a tapered shape, with a diameter on the orderof 300 microns at the top. This aperture may taper to a diameter ofabout 150 microns at the base where it meets the corresponding apertureof sort channel 20 of MEMS chip sorter 10.

FIG. 6 also illustrates the details of the mixing mechanism, which mayinclude a propeller 1150 mounted on a rotating shaft 1160. The shaft mayextend from the top surface of the cartridge 1135 through a bearingstructure which allows the shaft to rotate freely. If the propeller 1150contains magnetic components, the mixing action may be accomplished by arotating magnetic field external to the cartridge 1000. The appliedfield my drive the motion of the propeller, causing it to rotate on theshaft 1160. Alternatively, a mechanical coupling may engage a motorwhich then rotates the shaft 1160. Accordingly, the cell sorting system1 may include a disposable cartridge 1000, which may further include apropeller on a shaft, wherein the propeller 1150 is disposed in thesample reservoir 20. The propeller 1150 may comprise magnetic material,and which interacts with a variable magnetic field which turns thepropeller 1150 on its shaft 1160, thereby mixing the contents of thesample reservoir 20. Alternatively, the cell sorting system 1 mayinclude a propeller 1150 which is rotated by a mechanical coupling thatrotates the shaft 1160, and is driven by a motor.

The interposer 1400 may have passages formed therein, 1120, 1122 and1140, shown in FIG. 7, which may correspond to channels 120, 122 and 140shown in FIGS. 1 and 2. That is, passage 1120 may mate with passage 120on MEMS chip sorter 10, to provide a fluidic pathway from samplereservoir 20 to MEMS chip sorter 10. Downstream of MEMS chip sorter 10,the interposer 1400 may provide a fluidic pathway from the movable valve110 to the sort reservoir 22 (in cartridge) via sort channel 122 (onchip) and 1122 (on interposer). Similarly, the interposer 1400 mayprovide a fluidic pathway from the movable valve 110 to the wastereservoir 40 (in cartridge) via waste channel 140 (on chip) and 1140 (oninterposer). In other words, the interposer 1400 may provide a sortfluid path between a sort reservoir 22 in the disposable cartridge 1000and the sort channel 122 in the silicon substrate, a waste fluid pathbetween a waste reservoir 40 in the disposable cartridge 1000 and thewaste channel 140, and a sample fluid path between the sample channel120 and a sample reservoir 20.

Another purpose of the interposer 1400 is to provide a collection regionfor possibly small volumes of sorted material. For example, since thetarget cells may be rare, such as stem cells, the volume of fluidcollected in the sort reservoir 22 may also be rather small, and inproportion to the frequency of target cells in the sample. Accordingly,volumes as low as a few microliters may be expected. The interposer 1400may provide a region into which the sorted effluent is siphoned, foreasy collection with a small pipette. This siphon region 1450 is shownin FIG. 7.

In particular, it should be noticed that the floor of siphon region 1450is at a lower elevation than the bottom of the sort channel 1122.Accordingly, fluid may flow as assisted by siphoning action and meniscusforces from the MEMS chip sorter 10 to the sort reservoir 22, from whichit can be retrieved by hypodermic needle or micropipette. This siphoningmay help offset the capillary forces that may occur from small volumeflow in the very small channels. Accordingly, the sort reservoir 22 mayfurther comprise a siphon structure that collects a smaller sort fluidvolume within the sort reservoir 22, wherein the smaller sort fluidvolume is less than about 10% of the total fluid volume of the sortreservoir 22.

Importantly, the sort channel 1122 may be made relatively short comparedto sample channel 1120 and waste channel 1140, so that the amount ofmaterial lost by adhesion to channel walls, for example, is minimized.

Also shown in the detail of FIG. 7 is a glue dam 1460, which will bedescribed next with respect to cartridge assembly.

As can be seen in FIG. 7, the sample channel 1122 may draw material fromthe very bottom of the sample reservoir 20. This may be important inmaximizing the yield, or percent of recovered material, from a givensample volume. In contrast, the waste channel 1140 may deliver thenon-target material to a point on the incline of the wall of the wastevoid or reservoir 40.

The interposer 1400 may be made from polycarbonate, polymethylmethacrylate (PMMA), or cyclic olefin polymer (COP), by injectionmolding, embossing, laser machining or 3D printing. The tolerances onthe passages 1420 shown in FIG. 6 in the interposer 1400 may be about+/−10 microns on a total diameter of about 100 to 400 microns. Thecorresponding passages 20 in the MEMS chip sorter 10 may be about 50 to150 microns. These passages 20 and 1420 may then be aligned as was shownin the insert to FIG. 6 to within about 10 microns. The interposer 1400is affixed to the silicon substrate by any convenient adhesive, such asglue, epoxy and cement. The MEMS chip sorter 10 may first be glued tothe interposer by seating it in the chip cavity 1470 shown in FIG. 9.The cavity 1470 may be formed sufficiently precisely that the passagesin MEMS chip sorter 10 roughly overlap the passages in interposer 1400.The allowed mismatch may be up to about 20 microns, easily achievable. Apick and place machine, well known in printed circuit boardmanufacturing, may be adequate for this task. The MEMS chip sorter 10may be glued in place within cavity 1470. Of course, the materials andmethods described here are exemplary only. Other materials, such asother plastics, and other corresponding methods may be used.

The interposer 1400 may then be installed in the cartridge base 1130with glue or cement, by locating the interposer 1400 locating holes 1410against corresponding posts in cartridge body 1000. Since this glue orcement will be required to be watertight, yet not interfere withpassages 1120, 1122 or 1140, some features may be formed as glue dams1460 around these channels, as shown in FIGS. 7 and 8. These glue dams1460 may serve to keep the liquid, uncured glue from entering the smallchannels 1120, 1122 and 1140. The features 1460 may be raised ridges ofplastic material which prevent the liquid from entering the channels orother depressions. In particular, glue may be injected into a port thatgives access to the interface between interposer 1400 and the remainderof cartridge body 1000. The glue will wick around this area but may bekept out of microfluidic passageways 1120, 1140 and 1122 by glue dams1460 that surround these passageways as shown in FIG. 7 and in theperspective drawing of FIG. 8. The glue dams reduce the thickness of theinterface between interposer 1400 and the remainder of cartridge body1000 from about 5 to 50 μm to 0.2 to 2 μm thereby creating a capillaryeffect that prevents the glue from flowing beyond the dam into themicrofluidic passageways. It should be understood that these dimensionsare exemplary only, and that such details will depend on the specificsof the application. Depending on the type of glue used, the liquid gluemay be cured by heat, pressure or exposure to UV radiation, for example.

FIG. 9 is a simplified perspective view of the obverse side of theinterposer 1400. This side includes the seating area 1470 for MEMS chipsorter 10. The MEMS chip sorter 10 may be glued or otherwise bondedagainst the features of seating area 1470

Exemplary dimensions for the interposer are 16 mm length, 6 mm width, 1mm height. The waste and sample reservoirs may be 2 mm in diameter. Thesample channel 1120, sort channel 1122 and waste channel 1140 may eachbe 300 microns in width. The height of the glue dams may be about 20microns.

Accordingly, a manufacturing process for the cartridge 1000 may include:

-   -   1) Glue MEMS chip sorter 10 to interposer 1400    -   2) Place interposer 1400 against cartridge 1000 locating pins    -   3) Press interposer 1400    -   4) Introduce glue to gaps between interposer 1400 and cartridge        1000    -   5) UV cure glue    -   6) Attach cartridge base 1130 to cartridge top 1135 by glue,        cement, or ultrasonic welding, for example    -   7) This invention relates to a cell sorting system using a        microfabricated, movable cell sorting mechanism.

It should be clear that steps 1-6 need not be executed in the ordershown. For example, the cartridge base 1130 may be attached to thecartridge top 1135 before attaching the MEMS chip 10 or interposer 1400.

In another embodiment of the cell sorting system, the disposablecartridge and/or the interposer further comprise at least onecalibration region to calibrate the interrogation means.

FIG. 10 is a simplified diagram of the interposer 1400 comprisingcalibration regions. The interposer 1400 may have areas 1480 that areused for calibration purposes. These areas 1480 may have particular,pre-defined optical or fluorescence properties that are used for thecalibration or compensation of an optical system. Therefore, the areas1480 can be made of one or more fluorescent material or one or morefluorescent material is added (printed) to these surfaces.Alternatively, microfluidic structures may be used to guide and storeone or more fluorescent liquids to these calibration areas. Theapplication of these fluorescent liquids may therefore take place at adifferent position on the interposer. FIG. 11 is a cross sectional viewof such a variant of the interposer 1400 with calibration areas 1480.The interposer may have a sample channel 1120 as described above, whichbrings the sample fluid to the MEMS chip sorter 10. The fluid may thenpass into the calibration areas 1480.

The calibration is meant to make sure suspended matter having equivalentfluorescent staining gets consistently detected across instruments andover time. This is achieved by measuring the intensity of a materialwith known fluorescence. Instead of using calibration particles in adifferent run, this may be achieved by illuminating and detectingfluorescence from the calibration area on or in the interposer withinthe cartridge. Calibration may be carried out any time, for exampleafter inserting the cartridge in the system before starting the sortingprocess, or during a sorting process in order to control or maintainsystem performance.

By calibration against a known fluorescing material, a predefined targetintensity can be adjusted by the system of the invention. Calibrationcan be performed several times in an iterative process and can befurther utilized to characterize and validate system performance.

The fluorescing material used for the calibration area 1480 are selectedthat they can be detected in at least, one, at best all fluorescencedetection channels used in the system and that their fluorescenceintensity (absolute brightness) is at least in the same order ofmagnitude as the fluorescence intensity (absolute brightness) thematerial to be processed. Especially suitable as fluorescing materialfor the calibration area are Coumarin-6, Nile-Red and/or Bodipy-650.

In another embodiment of the invention, the fluid channel geometry ofthe interposer is designed to avoid trapping of bubbles andagglomeration of cellular material. Therefore, the channel geometriesmay be optimized with respect to fluidic properties. These features mayminimize dead volumes, and avoid undercuts and rounded corners.Furthermore, the channel geometries at the interfaces to the chip and/orthe cartridge main body may be designed in way that channel diametersare always increasing in flow direction. These design elements may alsoprevent the agglomeration of cellular material inside the fluidicchannels.

Avoidance of trapped of bubbles can further be achieved by providing thechannels in the interposer with a small channel at the bottom of themain channel (“channel-in-channel”). The small channel may have 5 to 20%of either the depth and/or with of the channel it is located in. FIG. 8bshows by way of example the a small channel 1490 at the bottom of achannel of the interposer, such as channels 1120 and 1140. A gas bubbleblocking the main channel cannot enter the small channel due to surfacetension and leaves the small channel open for flow of liquid. Anothervariant of avoiding trapped bubbles is shown in FIG. 8c , where channel1122 leading to the siphon is shaped like a ramp leading into thesiphon, thus avoiding sharp edges. Accordingly, the interposer mayinclude least one channel having with an additional small channeldisposed at the bottom of the at least one channel, wherein said smallchannel has between about 5 to about 20% of a depth and a width of theat least one channel.

Another aspect of the system described above with respect to FIGS. 1, 2and 3 is the need for a precisely localized magnetic field which willactuate the small, MEMS chip sorter 10.

As described previously, the actuation mechanism in the system shown inFIG. 3 may be electromagnetic. Because the movable valve 110 is sosmall, it is important to have the flux-generating structure be precise,low power and efficient. Such a structure is shown in FIGS. 10a, 10b and10 c.

The external source of magnetic field lines (magnetic flux) may beprovided outside the MEMS chip sorter 10, as was shown in FIGS. 2 and 3.This source may be an electromagnet 400. The electromagnet 400 mayinclude a permeable core 470 around which coils 460 are wound. The coils460 and core 470 generate a magnetic field which exits the pole of themagnet at the tip 450, diverges, and returns to the opposite pole, as iswell known from elementary electromagnetism. In general, there may be atrade-off between fewer layers of coils 460 for more effective heatdissipation, or more layers for greater flux (Amp*turn), and thusgreater force and higher speed. In one embodiment, the coils 460 haveone layer, but the magnet body will often have at most three layers ofcoils 460. When the electromagnet 400 is brought into the vicinity ofthe movable valve 110, and the coil 460 is energized, the coil 460 andcore 470 generate lines of flux that diverge rapidly from the tip 450.Accordingly, the movable member 110 is generally drawn toward the tip450 of the electromagnet 400 as shown in FIG. 12a , because thepermeable material is drawn into areas of increasing flux density.

As shown in FIG. 12a , the magnet 470 may be given a tapered shape,which may tend to further concentrate the magnetic flux in the regionaround the tip 450. The angle of the taper may be, for example, betweenabout 0 and to about 30 degrees from vertical. The aspect ratio (lengthof taper/average width of taper) may be around 2/1 for example, but maybe designed in a broad range of shapes. In order to focus the flux atthe tip 450, however, it may be advantageous to have the diameter at thetip be less than the diameter at the base of the tapered shape. FIG. 12bis an enlarged view of the tip of electromagnet 400, showing the taperedshape of the tip, face-on. FIG. 12c is a perspective view of the taperedshape 450, coils 460 and magnet body 470.

Magnetic modeling suggests that a electromagnet tip of the approximatewidth of the permeable elements 116 in the MEMS chip sorter 10 isoptimal, with a height of approximately the same order of magnitude. Thebase size is then determined by the taper angle. In one embodiment, thebase of the tapered shape may have a length of 2 to 5 mm and a width of0.5 to 2 mm. The tip of the tapered shape may be smaller than the baseand have rectangular dimensions of about 1.0 mm×0.7 mm, or at leastabout 0.4 mm×0.2 mm. It should be understood that these dimensions areexemplary only, and that such details will depend on the specifics ofthe application.

While various details have been described in conjunction with theexemplary implementations outlined above, various alternatives,modifications, variations, improvements, and/or substantial equivalents,whether known or that are or may be presently unforeseen, may becomeapparent upon reviewing the foregoing disclosure. Furthermore, detailsrelated to the specific methods, dimensions, materials uses, shapes,fabrication techniques, etc. are intended to be illustrative only, andthe invention is not limited to such embodiments. Descriptors such astop, bottom, left, right, back front, etc. are arbitrary, as it shouldbe understood that the systems and methods may be performed in anyorientation. Accordingly, the exemplary implementations set forth above,are intended to be illustrative, not limiting.

What is claimed is:
 1. A cell sorting system for sorting targetparticles from a fluid sample, comprising: a cell sorting valvemicrofabricated on a surface of a silicon substrate, withmicrofabricated channels leading from the cell sorting valve, whereinthe cell sorting valve separates the target particles from non-targetmaterial; a disposable cartridge containing a sample reservoir, a sortreservoir and a waste reservoir; and an interposer that provides fluidcommunication between the microfabricated channels in the siliconsubstrate and the reservoirs in the disposable cartridge and wherein thecell sorting valve moves in a plane parallel to the surface, and directsthe target particles from the sample channel into the waste channel whenthe microfabricated cell sorting valve is in a first position, and whichdirects the non-target particles into the sort channel when in a secondposition, wherein the waste channel is substantially orthogonal to theplane of motion of the cell sorting valve.
 2. The cell sorting system ofclaim 1, wherein the cell sorting valve directs the target particlesfrom a sample channel into a sort channel formed in the siliconsubstrate and the non-target material from the sample channel to a wastechannel also formed in the silicon substrate.
 3. A cell sorting systemfor sorting articles from a fluid sample, comprising: a cell sortingvalve microfabricated on a surface of a silicon substrate, withmicrofabricated channels leading from the cell sorting valve, whereinthe cell sorting valve separates the target particles from non-targetmaterial; a disposable cartridge containing a sample reservoir, a sortreservoir and a waste reservoir; and an interposer that provides fluidcommunication between the microfabricated channels in the siliconsubstrate and the reservoirs in the disposable cartridge, wherein thecell sorting valve moves in a plane parallel to the surface, and directsthe target particles from the sample channel into the waste channel whenthe microfabricated cell sorting valve is in a first position, and whichdirects the non-target particles into the sort channel when in a secondposition, wherein the sort channel and the waste channel aresubstantially antiparallel, and the sample channel and the waste channelare substantially orthogonal.
 4. The cell sorting system of claim 3,wherein the interposer provides a sort fluid path between a sortreservoir in the disposable cartridge and the sort channel in thesilicon substrate, a waste fluid path between a waste reservoir in thedisposable cartridge and the waste channel, and a sample fluid pathbetween the sample channel and a sample reservoir.
 5. The cell sortingsystem of claim 4, wherein the sort reservoir further comprises a siphonstructure that collects a smaller sort fluid volume within the siphonstructure of the sort reservoir, wherein the smaller sort fluid volumeis less than about 10% of a total fluid volume of the sort reservoir. 6.The cell sorting system of claim 4, wherein the sample reservoir furthercomprises a funnel-shaped feature formed in the wall of the samplereservoir, which collects a smaller volumes of sample fluid, wherein thesmaller volume of sample fluid is less that about 10% of a total fluidvolume of the sample reservoir.
 7. The cell sorting system of claim 4,wherein the waste reservoir further comprises a funnel-shaped featureformed in the wall of the waste reservoir, which collects a smallervolume of waste fluid, wherein the smaller volume of waste fluid is lessthat about 10% of a total fluid volume of the waste reservoir.
 8. Thecell sorting system of claim 3, wherein the interposer is affixed to thesilicon substrate by an adhesive, and installed in the disposablecartridge.
 9. The cell sorting system of claim 3, wherein the cellsorting valve further comprises: a permeable magnetic material inlaid inthe microfabricated cell sorting valve; and a first source of magneticflux external to the microfabricated cell sorting valve and substrate onwhich the cell sorting valve is formed, wherein the magnetic fluxinteracts with the inlaid permeable magnetic material to move themicrofabricated cell sorting valve, whereby the microfabricated cellsorting valve moves from the first position to the second position whenthe first source of magnetic flux is activated.
 10. The cell sortingsystem of claim 3, wherein when the cell sorting valve is in the firstposition, a passage between the sample channel and the waste channel isformed.
 11. The cell sorting system of claim 3, wherein when themicrofabricated cell sorting valve is in the second position, a passagebetween the sample channel and the sort channel is formed.
 12. The cellsorting system of claim 11, wherein the cell sorting valve moves fromthe first position to the second position in response to the applicationof a force.
 13. The cell sorting system of claim 12, wherein the forceis at least one of mechanical, electrostatic, magnetostatic,piezoelectric and electromagnetic.
 14. The cell sorting system of claim3, wherein the disposable cartridge further includes a propeller on ashaft wherein the propeller is disposed in the sample reservoir.
 15. Thecell sorting system of claim 14, wherein the propeller comprisesmagnetic material, and interacts with a variable magnetic field whichturns the propeller on its shaft, thereby mixing the contents of thesample reservoir.
 16. The cell sorting system of claim 14, wherein thepropeller is rotated by a mechanical coupling that rotates the shaft,and is driven by a motor.
 17. The cell sorting system of claim 3,further comprising: an interrogation means disposed in the samplechannel, wherein the interrogation means distinguishes target particlesfrom non-target materials in the sample stream.
 18. The cell sortingsystem of claim 17, wherein the interrogation means comprises a laser ina laser-based, induced fluorescence system, wherein a fluorescent tag isaffixed to the target particle, and emits a fluorescent signal whenirradiated by the laser.
 19. The cell sorting system of claim 3, whereinthe disposable cartridge is configured to be accepted into the cellsorting system on a positionable stage, on which it can be positionedwith respect to at least one laser source, and at least one opticaldetector.
 20. The cell sorting system of claim 19, wherein the cellsorting system further includes a computer which is in communicationwith the at least one laser source, at least one optical source and thecell sorting valve, to separate the target particles from the non-targetmaterial.
 21. The cell sorting system of claim 17, wherein thedisposable cartridge further comprises at least one calibration regionto calibrate the interrogation means.
 22. The cell sorting system ofclaim 17, wherein the interposer further comprises at least onecalibration region to calibrate the interrogation means.
 23. A cellsorting system for sorting target particles from a fluid sample,comprising: a cell sorting valve microfabricated on a surface of asilicon substrate, with microfabricated channels leading from the cellsorting valve, wherein the cell sorting valve separates the targetparticles from non-target material; a disposable cartridge containing asample reservoir, a sort reservoir and a waste reservoir; and aninterposer that provides fluid communication between the microfabricatedchannels in the silicon substrate and the reservoirs in the disposablecartridge, wherein the interposer comprises at least one channel havingan additional small channel disposed at the bottom of the at least onechannel, wherein said small channel has between about 5 to 20% of adepth and a width of the at least one channel.